Radhakrishnan I, Patel D J
Department of Biochemistry and Molecular Biophysics, College of Physicians and Surgeons, Columbia University, New York NY 10032.
Structure. 1993 Oct 15;1(2):135-52. doi: 10.1016/0969-2126(93)90028-f.
Oligonucleotide-directed triple helix formation allows sequence specific recognition of double helical DNA. This powerful approach has been used to inhibit gene transcription in vitro and to mediate single site specific cleavage of a human chromosome.
Using a combined NMR and molecular dynamics approach (including relaxation matrix refinement), we have determined the solution structure of an intramolecular purine.purine.pyrimidine (R.RY) DNA triplex containing guanines and thymines in the third strand to high resolution. Our studies define the G.GC and T.AT base triple pairing alignments in the R.RY triplex and identify the structural discontinuities in the third strand associated with the non-isomorphism of the base triples. The 5'-d(TpG)-3' base steps exhibit a pronounced increase in axial rise and reduction in helical twist, while the reverse is observed, to a lesser extent at 5'-d(GpT)-3' steps. A third groove is formed between the purine-rich third strand and the pyrimidine strand. It is wider and deeper than the other two grooves.
Our structure of the R.RY DNA triplex will be important in the design of oligonucleotide probes with enhanced specificity and affinity for targeting in the genome. The third groove presents a potential target for binding additional ligands.
寡核苷酸定向三链螺旋形成可实现对双链DNA的序列特异性识别。这种强大的方法已被用于体外抑制基因转录以及介导人类染色体的单一位点特异性切割。
使用核磁共振（NMR）和分子动力学相结合的方法（包括弛豫矩阵精修），我们已高分辨率地确定了第三条链中含有鸟嘌呤和胸腺嘧啶的分子内嘌呤·嘌呤·嘧啶（R.RY）DNA三链体的溶液结构。我们的研究确定了R.RY三链体中的G.GC和T.AT碱基三联配对排列，并识别出与碱基三联体非同构相关的第三条链中的结构不连续性。5'-d（TpG）-3'碱基步的轴向上升明显增加，螺旋扭转减小，而在5'-d（GpT）-3'步则观察到相反情况，但程度较小。在富含嘌呤的第三条链和嘧啶链之间形成了第三条凹槽。它比其他两条凹槽更宽更深。
我们的R.RY DNA三链体结构对于设计对基因组靶向具有更高特异性和亲和力的寡核苷酸探针至关重要。第三条凹槽为结合额外配体提供了潜在靶点。
